Protease is a proteolytic enzyme and there are known various kinds of protease, including those showing an antiinflammatory and antihydropic effect, those showing a pus dissolving action and those showing a blood coagulation inhibiting action. These proteases are applied to the preparation of pharmaceuticals. With such kinds of proteases, especially those which are administered to man through intravenous injection exclusively, such as urokinase, safety must be guaranteed in their application, and thus it is an essential requirement to obtain high-purity proteases.
Various techniques and methods have been developed for the purification of the crude preparations of proteases. Recently, the affinity chromatographic techniques using an adsorbent prepared from a substance having a high affinity for the protein to be purified and combined with a water-insoluble support as a ligand are applied to the purification of proteases. For instance, the following methods have been proposed for the purification of urokinase:
(1) A basic amino acid such as lysine or arginine or a derivative thereof is used as the ligand (Japanese Patent Publication No. 44193/1976, and Japanese Patent Laid-Open Nos. 20596/1976, 95183/1976 and 35481 to 35483/1976).
(2) A urokinase inhibitor contained in placental tissues, etc., is used as the ligand (Japanese Patent Publication No. 20597/1976).
In the method (1), however, the substance used as the ligand has no satisfactory affinity for urokinase and it is hardly possible to specifically adsorb urokinase from a solution with a high salt concentration. The method (2) is rather impractical because, in this method, an inhibitory substance found only in a small quantity in the animal tissues is used as the ligand.